CD8-LYT-2 RECOGNITION OF THE CLASS I ALPHA 3 DOMAIN

I类反应性T细胞表面的CD8分子最初是 据信起到辅助分子的作用，增加了对 第一类MHC的TCR。最近的证据表明，CD8分子可能 与TCR结合以获得抗原，从而在信号中发挥作用 转导和激活。现在看来，CD8受体也是 参与I类反应性T的阳性和阴性选择 胸腺成熟过程中的细胞。校长最近的观察 CD8与I类alpha3结构域中残基结合的调查者是 首次鉴定CD8识别的单态行列式。的 在这些研究中，最重要的是使用DD Alpha3结构域 突变体Ddlys，已经失去了被CD8识别的能力。这个 这项提案的主要目的是探讨有关 CD8在T细胞识别中的作用 CD8分子I类特异性CTL：I类识别残基 Alpha3域。鉴于之前的研究涉及异质 CTL的种群新的研究将使用克隆种群 CD8依赖和CD8非依赖的CTL。这些克隆人将被分析 它们裂解表达Ddlys和其他α3结构域的细胞的能力 对于突变体以及针对α3结构域和CD8的抗体 抑制它们的功能。它们还将用于细胞：细胞结合分析 确定它们对CD8细胞结合的依赖性以及 细胞结合的α3结构域突变体。额外的α3结构域突变体 通过定点突变获得的基因由T.Potter博士提供。 这些新的突变体将在各种测试中进行比较，并将确定 CD8识别的I类配体的范围。此外，CTL 对I类α3结构域突变的反应将被分析为 确定CD8识别是否涉及其他残基。 最后，CD8参与了CD8的生成和效应功能。 将分析MHC限制的CTL反应。因此，有几个突出的 CD8的特性将用多种免疫学方法进行研究 检测和一组独特的I类突变位点。

The CD8 molecule on the surface of class I reactive T cells was initially believed to function as an accessory molecule, increasing the affinity of the TCR for class I MHC. Recent evidence suggests that CD8 molecules may associate with the TCR for antigen and thereby function in signal transduction and activation. It appears now that the CD8 receptor is also involved in both positive and negative selection of class I reactive T cells during thymic maturation. The recent observation by the principal investigator the CD8 binds to residues in the class I alpha3 domain was the first identification of the monomorphic determinant recognized by CD8. Of paramount importance in these studies was the use of the Dd alpha 3 domain mutant, Ddlys, that had lost the ability to be recognized by CD8. The primary objectives of this proposal are to probe questions concerning the role of CD8 in T cell recognition and to extend the observation that the CD8 molecule o class I-specific CTL: recognized residues in the class I alpha3 domain. Whereas the previous studies involved heterogeneous populations of CTL the new investigations will employ clonal populations of CD8-dependent and CD8-independent CTL. These clones will be analyzed for their ability to lyse cells expressing Ddlys and other alpha3 domain mutants and for the ability of antibodies to the alpha3 domain and CD8 to inhibit their function. They will also be used in cell:cell binding assays to determine their dependency on CD8 for cell binding and the effect of alpha3 domain mutants of cell binding. Additional alpha3 domain mutants obtained by site directed mutagenesis are being provided by Dr. T. Potter. These new mutants will be compared in a variety of assays and will define the extent of the class I ligand recognized by CD8. In addition, the CTL response against the class I alpha3 domain mutants will be analyzed to determine if additional residues are involved in recognition by CD8. Finally, the involvement of CD8 in the generation and effector function of MHC-restricted CTL responses will be analyzed. Thus several salient properties of CD8 will be investigated using a variety of immunologic assays and a unique collection of site-specific class I mutants.